DESCRIPTION: Binding of dopamine D2 PET and SPECT radioligands including [C-11] raclopride and [I-123]IBZM in vivo is affected by competition with endogenous dopamine. This allows changes in synaptic cleft levels of dopamine to be detected in human subjects, via corresponding increases or decreases in radioligand binding. If the factors which affect competition between exogenous radioligands and neurotransmitters were better understood, quantitative measures of the average concentrations of dopamine in the synaptic cleft could be obtained in conditions such as substance abuse and schizophrenia. As yet, however, the information needed to relate changes in D2 radioligand binding to changes in dopamine is lacking. Based on preliminary studies, it is proposed to further develop a superfused brain slice preparation, in which synaptic cleft neurotransmitter levels can be manipulated by controlling the rate of external electrical stimulation to the slice, while radioligands and pharmacological agents can be administered under very controlled conditions, including maintenance of radioligand concentrations at defined and constant values. The slices contain living cells with apparently normal synaptic connections, so that the local factors affecting competition in vivo are believed to be retained. The preparation represents a model of tomographically isolated brain tissue. The proposed experiment will thoroughly characterize the binding of several D2 radioligands to the slices, in terms of the kinetics of association and dissociation, and equilibrium binding parameters. Effects of electrical stimulation of the slices, of the dopamine reuptake blocker cocaine, and of the dopamine releasing drug amphetamine on radioligand binding will be evaluated. The influences of experimental reduction in D2 receptor density, of radiotracer affinity and of equilibrium versus non-equilibrium radioligand binding will also be examined. In addition to radioligand binding, a functional assay of dopamine concentration, the inhibition of electrically evoked acetylcholine release, will be employed. The average dopamine concentrations corresponding to a range of stimulation frequencies will be assessed using equilibrium radioligand binding models, and some of the anticipated conclusions of the studies will be tested in an animal model. The proposed studies are important both for the design and interpretation of PET/SPECT studies with currently available radioligands, and for elucidation of the factors necessary for design of optimum radiotracers for neurotransmitter competition experiments.